Genome mining in Amycolatopsis balhimycina for ferredoxins capable of supporting cytochrome P450 enzymes involved in glycopeptide antibiotic biosynthesis

Ferredoxins are required to supply electrons to the cytochrome P450 enzymes involved in cross-linking reactions during the biosynthesis of the glycopeptide antibiotics balhimycin and vancomycin. However, the biosynthetic gene clusters for these antibiotics contain no ferredoxin- or ferredoxin reductase-like genes. In a search for potential ferredoxin partners for these P450s, here, we report an in silico analysis of the draft genome sequence of the balhimycin producer Amycolatopsis balhimycina, which revealed 11 putative Fe-S-containing ferredoxin genes. We show that two members (balFd-V and balFd-VII), produced as native-like holo-[3Fe-4S] ferredoxins in Escherichia coli, could supply electrons to the P450 OxyB (CYP165B) from both A. balhimycina and the vancomycin producer Amycolatopsis orientalis, and support in vitro turnover of peptidyl carrier protein-bound peptide substrates into monocyclic cross-linked products. These results show that ferredoxins encoded in the antibiotic-producing strain can act in a degenerate manner in supporting the catalytic functions of glycopeptide biosynthetic P450 enzymes from the same as well as heterologous gene clusters.